Synthesis of 3′-O-phosphonomethyl nucleosides with an adenine base moiety

Article abstract of DOI:10.1016/j.tet.2007.01.032

A synthetic scheme has been developed for the synthesis of 2′-deoxythreose phosphonate nucleosides from β-hydroxy-γ-butyrolactone and of 2′-azido erythrose phosphonate nucleosides from dihydroxydihydrofuran-1-one. In addition several α-l-arabinofuranose p

Full text of DOI:10.1016/j.tet.2007.01.032

Tetrahedron 63 (2007) 2634–2646
Synthesis of 30-O-phosphonomethyl nucleosides with an adenine
base moiety
a
Dolores Vina, Tongfei Wu, Marleen Renders, Genevieve Laflamme and Piet Herdewijn
a
a
b
a,
*
ꢀ
˜
aLaboratory of Medicinal Chemistry, Rega Institute for Medical Research, K.U. Leuven,
Minderbroedersstraat 10, B-3000 Leuven, Belgium
bGilead Sciences, 333 Lakeside Drive, Foster City, CA 94404, USA
Received 23 November 2006; revised 12 January 2007; accepted 17 January 2007
Available online 20 January 2007
Abstract—A synthetic scheme has been developed for the synthesis of 20-deoxythreose phosphonate nucleosides from b-hydroxy-g-
butyrolactone and of 20-azido erythrose phosphonate nucleosides from dihydroxydihydrofuran-1-one. In addition several a-L-arabinofuranose
phosphonate nucleosides were synthesized starting from protected a-D-galactofuranose. Unfortunately, none of the synthesized compounds
show activity in an HIV-1 assay. One of these compounds, a locked phosphonate nucleoside, was evaluated (as diphosphate) for its potential to
be incorporated into DNA using HIV-1 reverse transcriptase.
Ó 2007 Elsevier Ltd. All rights reserved.
1. Introduction
bridge (5). The reason for synthesizing these compounds is
to map the substitution landscape (especially in the 20- and
40-position) as a function of antiviral activity. The triphos-
phate of the locked phosphonate nucleoside was synthesized
and tested as potential substrate for HIV-1 reverse transcrip-
tase of PMDTA.
One of the advantages of the antiviral phosphonate nucleo-
sides that are currently on the market for the treatment
of HIV infections is that they need to be given only once
daily. Limitations of the currently used phosphonate drug
(TenofovirÒ), that need to be surpassed, are the long-term
side effects, the emergence of resistance, and the presence
of drug–drug interactions.1 This research has led to the dis-
covery of three types of phosphonate nucleosides with an
adenine base moiety. The D-d4AP nucleoside was described
by Kim et al. in 1991.2 PMDTA (Fig. 1) was described as
a selective anti-HIV-1/HIV-2 phosphonate nucleoside.3
Both compounds demonstrated an anti-HIV activity (EC50)
of 2.0–3.0 mM against HIV-1. 20Fd4AP1 is a somewhat lesser
active compound (EC50: 12 mM) but with an excellent resis-
tance profile (Fig. 1). In order to investigate the importance
of the stereochemistry in the 10- and 30-position of PMDTA,
we have now synthesized the isomeric nucleosides 1a and 1b
(Fig. 2). PMDTA has a phosphonomethyl moiety at the 30-
position of the furanose ring and no substituent at the 40-
position.3 The absence of a 40-hydroxymethyl group avoids
problems of steric hindrance during phosphorylation reac-
tions by kinases. To further study the influence of different
substituents on the anti-HIV activity, we have synthesized
20- and 40-modified analogs of PMDTA (2–4) as well as a
locked phosphonate nucleoside with a 40,20-O-a-methylene
2. Results and discussion
The nucleosides 1a and 1b were synthesized starting from
(S)-3-hydroxy-g-butyrolactone (Scheme 1). The hydroxyl
group in 3-position of (S)-3-hydroxy-g-butyrolactone is pro-
tected by benzoylation4 and the lactone function is reduced
using Dibal-H in THF.5 The anomeric hydroxyl group of
compound 7 is protected with acetic anhydride in pyridine.
The nucleobase (N6-benzoyladenine) is introduced using
SnCl4 as Lewis catalyst,6 giving a mixture of compounds
9a and 9b with the base moiety in b- and a-configuration, re-
spectively. The benzoyl protecting groups of 9a and 9b are
removed using satd ammonia in methanol. Finally, the phos-
phonate function is introduced on compounds 10a and 10b7
using the triflate of diisopropylphosphonomethyl alcohol
and NaH in THF. Hydrolysis of the phosphonate ester func-
tion of 11a and 11b is carried out with TMSI at 0 ꢀC.8 After
purification by silica gel chromatography, Sephadex-DEAE
A-25 resin and Dowex-sodium ion exchange resin, nucleo-
side phosphonates 1a and 1b were obtained.
Keywords: Nucleosides; Phosphonates; HIV; Threose; Erythrose.
* Corresponding author. Tel.: +32 16 337387; fax: +32 16 337340; e-mail:
piet.herdewijn@rega.kuleuven.be
Nucleoside 2 was synthesized starting from (R,R)-2,3-dihy-
droxydihydrofuran-1-one (Scheme 2). Compound 13, with
0040–4020/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2007.01.032

D. Vin˜a et al. / Tetrahedron 63 (2007) 2634–2646
2635
NH2
NH2
NH2
N
N
N
N
N
N
N
O
N
N
O
O
HO
P
O
N
O
HO
O
P
OH
N
O
N
O
OH
O
HO
P
OH
F
D-d4AP
PMDTA
D-2'Fd4AP
Figure 1. Structure of representative anti-HIV phosphonate nucleosides.
a TBDMS group at the 3-position, is a minor compound
when selective protection of the 2-position of 12 is carried
out with tertbutyldimethylsilyl chloride.3 We realize that
this multistep synthesis uses a compound, obtained in low
yield, as starting material. However, no attempts were
made to improve this yield, at this moment. The hydroxyl
group in 2-position of compound 13 is protected with
benzoyl chloride in pyridine. The lactone function is reduced
to hemiketal using Dibal-H in THF. The anomeric hydroxyl
group of compound 15 is protected with acetic anhydride in
pyridine. The nucleobase (adenine) is introduced using
SnCl4 as Lewis catalyst. The presence of a 2-O-benzoyl
group of compound 15 allows selective introduction of the
base moiety in the a-configuration. Then the benzoyl pro-
tecting group is removed using satd ammonia in methanol.
The introduction of the azido function into the 20-position
of compound 17 is performed through a nucleophilic substi-
tution of the triflate-activated 20-hydroxyl group with lithium
azide.9 The silyl protecting group is removed and the phos-
phonate function is then introduced in this position using the
triflate of diisopropylphosphonomethyl alcohol and NaH in
THF. Hydrolysis of the phosphonate ester function of com-
pound 20 is carried out with TMSI at 0 ꢀC. After purification
by silica gel chromatography, Sephadex-DEAE A-25 resin
and Dowex-sodium ion exchange resin, nucleoside phospho-
nate 2 was obtained.
The nucleosides 3–5 (Scheme 3) were synthesized starting
from 1,2:5,6-di-O-isopropylidene-a-D-galactofuranose 21,
which was obtained according to the procedure described
previously.10 The phosphonate function is introduced on
O
O
O
O
O
a
O
OH
OAc
c
b
7
HO
BzO
BzO
BzO
6
8
d
NHBz
NHBz
N
N
N
N
N
N
N
O
e
O
N
+
9a
9b
BzO
BzO
NH2
e
N
N
R1
O
NH2
NH2
N
N
N
N
O
P
O
O
P
N
N
N
R2
N
N3
O
HO
O
HO
O
O
N
OH
OH
N
O
1a
1b
R
R
R
R
1 = adenin-9-yl
2
10a
10b
2 = H
HO
HO
NH2
1 = H
NH2
2 = adenin-9-yl
f
N
N
f
N
N
N
NH2
N
O
NH2
N
N
N
N
N
O
P
O
N
N
O
N
O
O
O
O
P
O
P
ONa
N
11a
11b
O
O
O
P
ONa
HO
N
O
NaO
O
NH2
NaO
O
NH2
OH
g
N
N
N
g
OH
Cl
N
N
N
O
3
4
N
O
N
O
P
NH2
O
HO
O
P
OH
O
OH
N
N
HO
N
1a
1b
O
P
ONa
N
O
Scheme 1. Synthesis of two stereoisomers of PMDTA. Reagents and condi-
tions: (a) BzCl, pyridine, 89%; (b) Dibal-H, THF, 75%; (c) Ac2O, Et3N,
DMAP, 83%; (d) SnCl4, MeCN, N6-benzoyladenine, 9a: 19%, 9b: 33%;
(e) NH3/MeOH, 86%; (f) trifluoromethanesulfonate of diisopropylphospho-
nomethanol, NaH, THF, 52%; (g) (1) TMSI, DCM; (2) Sephadex-DEAE,
Dowex-Na+, 45%.
NaO
O
O
5
Figure 2. Structure of the target nucleosides.

2636
D. Vin˜a et al. / Tetrahedron 63 (2007) 2634–2646
O
O
O
O
O
O
O
OAc
c
a
b
HO
OH
TBDMSO
OH
TBDMSO
OBz
TBDMSO
OBz
15
12
14
13
d
NH2
NH2
NH2
N
N
N
N
N
N
N
N
f
e
O
O
O
N
N
N
N
TBDMSO
OBz
TBDMSO
TBDMSO
N3
OH
16
18
17
g
NH2
NH2
NH2
N
N
N
N
N
N
N
N
O
N
O
N
h
O
N
i
N
O
O
O
N3
O
P
O
HO
N3
O
N3
HO
P
OH
20
2
19
Scheme 2. Synthesis of the 20-deoxy-20-azidophosphonate nucleoside. Reagents and conditions: (a) TBDMSCl, imidazole, MeCN, 10%; (b) BzCl, pyridine,
85%; (c) (1) Dibal-H, THF, 94%; (2) Ac2O, Et3N, 58%; (d) SnCl4, MeCN, adenine, 39%; (e) NH3/MeOH, 99%; (f) (1) trifluoromethanesulfonyl chloride,
CH2Cl2; (2) LiN3/DMF, 37%; (g) TBAF/THF, 86%; (h) trifluoromethanesulfonate of diisopropylphosphonomethanol, NaH, THF, 74%; (i) (1) TMSI, DCM;
(2) Sephadex-DEAE, Dowex-Na+, 41%.
21 using the triflate of diisopropylphosphonomethyl alcohol
and NaH in THF,3 followed by oxidative degradation of
compound 22 to the aldehyde derivative 23 using H5IO6
and NaIO4.11 Wittig olefination of compound 23 with (chloro-
methyl)triphenylphosphonium chloride affords the chloro-
vinyl derivative 24.12 The isopropylidene protecting group
of 24 is removed and replaced by benzoyl protecting groups.
N6-Benzoyladenine is introduced using SnCl4 as Lewis
catalyst. Removal of the benzoyl protecting groups with
ammonia in methanol affords compound 27.
2.1. Biology
Compounds 1a, 1b, and 2–5 were evaluated as inhibitor of
HIV-1 replication in MT-2 cells. None of the compounds
shows activity at the highest concentration tested (200 mM).
Because the phosphonate with the locked conformation lacks
a hydroxyl group on the 20-position, it is important to check
whether this phosphonate nucleoside 35 can possibly act as
a chain terminator. In order to behave as a chain terminator,
the nucleotide analog should be incorporated into a growing
DNA or RNA strand and prevent further elongation.
After reduction of the aldehyde group of compound 23 to the
free hydroxyl group with NaBH4,13 the free hydroxyl group
of 28 was protected with a benzoyl group. The isopropyl-
idene protection group of 29 is removed and replaced by
benzoyl protecting groups. The presence of a 2-O-benzoyl
group allows selective introduction of the base moiety N6-
benzoyladenine using SnCl4 as Lewis catalyst in the a-con-
figuration. Removal of the benzoyl protecting groups with
ammonia in methanol afforded 32. After introduction of
a mesyl group on 32, locked nucleoside 34 is obtained by
intramolecular substitution using NaH.14
The ability of the phosphonate nucleoside 35 to terminate
DNA synthesis was investigated by a single nucleotide
incorporation assay using its diphosphate as potential
substrate.
Even with a high concentration of diphosphorylphosphonate
nucleoside (400 mM) and an enzyme (HIV reverse transcrip-
tion) concentration of 1.44 U/mL, only weak incorporation
(18% of the intact primer was elongated) could be seen.
Therefore, only in extreme circumstances the phosphonate
nucleoside will be able to act as a chain terminator.
The diisopropyl protected nucleoside phosphonates (27, 32,
and 34) were hydrolyzed with TMSI. After purification by
silica gel chromatography, Sephadex-DEAE A-25 resin and
Dowex-sodium ion exchange resin, nucleoside phosphonates
as sodium salts (3–5) were obtained. We synthesized the
diphosphate 35 of compound 5 (Fig. 3) to evaluate the poten-
tial of a locked nucleoside to function as chain terminator in
a polymerase-catalyzed reaction. This diphosphate was syn-
thesized using carbonyldiimidazole as coupling agent.
3. Conclusion
The influence of stereochemistry on the anti-HIVactivity of
tetrahydrofuran nucleosides with a phosphonomethoxy sub-
stituent in the 30-position has been investigated. Likewise,
a L-erythrose nucleoside with a 20-azido substituent and three

D. Vin˜a et al. / Tetrahedron 63 (2007) 2634–2646
2637
O
P
OPri
O
O
A
PriO
PriO
P
O
b
O
HO
O
O
O
a
OPri
99%
84%
O
O
O
O
O
O
O
O
O
O
21
22
23
NH2
c
N
N
N
O
O
O
N
PriO
O
O
PriO
PriO
1) d
2) e
O
P
OPri
O
P
OPri
O
P
OPri
O
OBz
f
73%
O
Cl
40% two steps Cl
OBz
OBz
O
Cl
26
25
24
g
88%
g
NH2
NH2
N
N
N
N
N
O
O
N
N
N
PriO
O
P
O
O
O
NaO
P
h
OPri
ONa
20%
OH
OH
Cl
Cl
3
27
B
23
a
99%
O
P
O
P
OPri
BzO
O
P
R2
PriO
PriO
PriO
O
O
O
O
O
O
1) c
b
OPri
BzO
OPri
R1
OBz
2) d
72%
O
68%
O
HO
O
O
30a R1=H, R2=OBz
30b R1=OBz, R2=H
29
28
NHBz
N
NH2
NH2
e
61%
O
N
N
N
N
N
N
N
O
P
O
O
N
N
f
PriO
O
O
N
PriO
O
NaO
h
P
N
O
P
OPri
O
g
OPri
ONa
87%
38%
OBz
OH
BzO
OH
HO
HO
31
32
4
NH2
NH2
NH2
N
48%
N
N
N
N
N
N
N
O
P
O
O
N
N
PriO
O
N
N
g
O
O
O
O
PriO
P
OPri
NaO
P
O
i
OPri
ONa
44%
83%
O
OH
O
MsO
5
34
33
Scheme 3. Synthesis of the 40-branched phosphonate nucleosides. (A) Reagents and conditions: (a) trifluoromethanesulfonate of diisopropylphosphonomethanol,
NaH, THF, ꢁ78 ꢀC, room temperature; (b) NaIO4, H5IO6, EtOAc, room temperature; (c) ClCH2P(C6H5)3Cl, n-BuLi, THF, ꢁ78 ꢀC; (d) TFA/H2O, room temper-
ature; (e)BzCl, pyridine, 0 ꢀC, roomtemperature; (f) SnCl4, MeCN, 0 ꢀC, roomtemperature; (g)satdNH3 inMeOH, roomtemperature;(h)(1)TMSI, DCM, 0 ꢀC;
(2)Sephadex-DEAE; (3)Dowex-Na+. (B)Reagentsandconditions: (a)NaBH4, MeOH, roomtemperature;(b)ClCH2P(C6H5)3Cl, n-BuLi, THF, ꢁ78 ꢀC;(c)TFA/
H2O, room temperature; (d) BzCl, pyridine, 0 ꢀC, room temperature; (e) SnCl4, MeCN, 0 ꢀC, room temperature; (f) satd NH3 in MeOH, room temperature; (g) (1)
TMSI, DCM, 0 ꢀC; (2) Sephadex-DEAE; (3) Dowex-Na+; (h) MsCl, pyridine, 0 ꢀC, room temperature; (i) NaH, THF, ꢁ78 ꢀC, room temperature.
examples of 40-branched phosphonate nucleosides have been
synthesized, but none of them show activity against HIV-1.
NH2
N
N
The diphosphate analog 35 was synthesized to test its ability
to function as chain terminator. Because of the flexibility of
the phosphonomethoxy moiety connected at the 30-position
of the sugar moiety, hypothesis about the conformational
preference of the sugar moiety for incorporation into
DNA (to predict the potential outcome of this incorporation
assay) was not taken into consideration. The phosphonate
nucleoside 35 could act as a chain terminator, only in
O
O
P
O
N
N
O
O
O
HO
O
P
P
OH
OH
OH
O
35
Figure 3. Structure of the diphosphate of the locked phosphonate nucleo-
side 5.

2638
D. Vin˜a et al. / Tetrahedron 63 (2007) 2634–2646
extreme conditions. Given its low potency, it is unlikely that
its parent phosphonate nucleoside 5 will be able to prevent
the virus to replicate under physiological circumstances
(even if sufficient amount of the diphosphate 35 could be
generated into the cell by the action of kinases).
to afford 7 (590 mg, 2.8 mmol) as a colorless oil in 75%
yield.
1H NMR (200 MHz, CDCl3) dH 2.30–2.40 (m, 2H, C(20)H2),
4.05–4.36 (m, 2H, C(40)H2), 5.30–5.82 (m, 2H,
C(10)H+C(30)H), 7.40–7.62 (m, 3H, Ar-H), 8.00–8.07 (m,
2H, Ar-H); 13C NMR (200 MHz, CDCl3) dC 39.67, 40.30
(C-20)71.38, 72.63 (C-30 or -40), 73.94, 74.97 (C-40 or -30),
98.37, 98.43 (C-10), 128.44, 128.53 (Ar-C), 129.69 (Ar-C),
133.27, 133.39 (Ar-C), 166.44 (Bz-CO); Exact mass calcd
for C11H12O4Na [M+Na]+ 231.0633, found 231.0629.
4. Experimental section
4.1. General
For all reactions, analytical grade solvents were used. All
moisture sensitive reactions were carried out in oven-dried
glassware (135 ꢀC) under a nitrogen atmosphere. Anhydrous
THF was refluxed over sodium/benzophenone and distilled.
A Varian Unity 500 MHz spectrometer and a 200 MHz
Varian Gemini apparatus were used for 1H NMR and
13C NMR. Exact mass measurements were performed on
a quadrupole time-of-flight mass spectrometer (Q-Tof-2,
Micromass, Manchester, UK) equipped with a standard elec-
trospray-ionization (ESI) interface; samples were infused in
i-PrOH/H2O 1:1 at 3 mL/min. Precoated aluminum sheets
(Fluka Silica gel/TLC-cards, 254 nm) were used for TLC;
the spots were examined with UV light. Column chromato-
graphy was performed on ICN silica gel 63-200. NMR sig-
nals of sugar protons and carbons are indicated with a
prime, signals of base protons and carbons are given without
a prime.
4.1.3. (S)-1-a,b-O-Acetyl-3-benzoyloxy-4-hydroxybuta-
nal (lactol) (8). To a solution of 7 (150 mg, 0.72 mmol) in
6.5 mL Et3N were added dropwise (CH3CO)2O (0.34 mL)
and DMAP (8.8 mg, 0.072 mmol) at 0 ꢀC. The reaction mix-
ture was warmed to room temperature and stirred for 3 h,
then concentrated in vacuo, and the residue was purified
by chromatography on a silica gel column (n-hexane/
EtOAc¼9:1) to give compounds 8a (117 mg, 0.47 mmol)
as colorless oil in 65% yield and 8b (32 mg, 0.13 mmol)
as colorless oil in 18% yield. It was not identified at this
stage which compound represents which isomer (a or b),
but the mixture of 8a and 8b was used in the condensation
reaction with the protected nucleobase.
Compound 8a: 1H NMR (200 MHz, CDCl3) dH 2.07 (s, 3H,
CH3), 2.48–2.52 (m, 2H, C(20)H2), 4.14 (dd, J1¼10.6 Hz,
J2¼2.2 Hz, 1H, C(40)Hb), 4.26 (dd, J1¼10.6 Hz, J2¼
4.2 Hz, 1H, C(40)Ha), 5.61–5.65 (m, 1H, C(30)H), 6.49
(t, J¼4.0 Hz, 1H, C(10)H), 7.40–8.05 (m, 5H, Ar-H).
Compound 8b: 1H NMR (200 MHz, CDCl3) dH 2.07
(s, 3H, CH3), 2.41–2.58 (m, 2H, C(20)H2), 4.22
(dd, J1¼10.6 Hz, J2¼2.6 Hz, 1H, C(40)Hb), 4.36 (dd,
J1¼10.6 Hz, J2¼5.2 Hz, 1H, C(40)Ha), 5.61–5.65 (m, 1H,
C(30)H), 6.40 (d, J¼4.4 Hz, 1H, C(10)H), 7.43–7.60
(m, 3H, Ar-H), 8.05–8.09 (m, 2H, Ar-H).
4.1.1. (S)-3-Benzoyloxy-g-butyrolactone (6). To a solution
of (S)-3-hydroxy-g-butyrolactone (1.0 g, 9.8 mmol) in
25 mL pyridine was added dropwise BzCl (1.4 mL,
12.2 mmol) at 0 ꢀC. The reaction mixture was warmed to
room temperature and stirred overnight. The reaction mix-
ture was concentrated and coevaporated with toluene two
times in vacuo. The residue was partitioned between H2O
(15 mL) and EtOAc (40 mL). The organic layer was washed
with water and brine, dried with Na2SO4, and concentrated
in vacuo. The residue was purified by chromatography on
a silica gel column (n-hexane/EtOAc¼8:1) to afford 6
(1.78 g, 8.6 mmol) as a white solid in 89% yield.
4.1.4. 1-b-(N6-Benzoyladenin-9-yl)-3-O-benzoyl-D-threo-
tetrafuranose (9a) and 1-a-(N6-benzoyladenin-9-yl)-3-
O-benzoyl-D-erythro-tetrafuranose (9b). To a mixture of
8 (210 mg, 0.84 mmol) and N6-benzoyladenine (405 mg,
1.62 mmol) in dry MeCN (30 mL) was dropwise added
SnCl4 (0.3 mL, 2.5 mmol) under N2 at 0 ꢀC. The reaction
mixture was warmed to room temperature and stirred for
1.5 h. Then the reaction was quenched with satd NaHCO3
and concentrated. The residue was partitioned between
H2O (20 mL) and EtOAc (100 mL). The organic layer was
washed with water and brine, and concentrated under vac-
uum. The residue was purified by chromatography on a silica
gel column (CH2Cl2/MeOH¼40:0.5) to afford a mixture of
b/a isomers (187 mg, 0.44 mmol) in 52% yield, which
were separated using preparative TLC and using CH2Cl2/
MeOH/40:1 (three times), as eluent.
1H NMR (200 MHz, CDCl3) dH 2.76–3.08 (m, 2H, C(20)H2),
4.58–4.70 (m, 2H, C(40)H2), 5.70–5.75 (m, 1H, C(30)H),
7.49–8.07 (m, 5H, Ar-H); 13C NMR (200 MHz, CDCl3) dC
33.21 (C-20), 68.88 (C-40), 71.73 (C-30), 127.24 (Ar-C),
128.43 (C-Ar), 132.43 (C-Ar), 169.05 (Bz-CO), C-10 was
obscured in noise peak; Exact mass calcd for C11H10O4Na
[M+Na]+ 229.0477, found 229.0435.
4.1.2. (S)-3-Benzoyl-4-hydroxybutanal (lactol) (7). To
a solution of 6 (0.780 g, 3.8 mmol) in 13 mL dry THF was
slowly dropwise added 1.0 M diisopropyl aluminum hydride
(4.7 mL, 4.7 mmol) in toluene at ꢁ78 ꢀC. The reaction mix-
ture was stirred at ꢁ78 ꢀC, and as soon as the starting mate-
rial was completely consumed (TLC, 1.5–2 h), methanol
(2 mL) was slowly added to quench the reaction. The cool-
ing bath was removed, 15 mL of a satd aq sodium potassium
tartrate solution and 25 mL of EtOAc were added, and the
mixture stirred vigorously for 3 h. The organic layer was
washed with water and brine, dried with Na2SO4, and
concentrated in vacuo. The residue was purified by chroma-
tography on a silica gel column (n-hexane/EtOAc¼8:2)
1
Compound 9a: H NMR (500 MHz, CDCl3) dH 2.84–2.89
(m, 1H, C(20)Hb), 3.23–3.28 (m, 1H, C(20)Ha), 4.29 (d,
J¼10.6 Hz, 1H, C(40)Hb), 4.63 (dd, J1¼4.3 Hz, J2¼
10.6 Hz, 1H, C(40)Ha), 5.92 (m, 1H, C(30)H), 6.50 (t,
J¼6.5 Hz, 1H, C(10)H), 7.45–7.60 (m, 6H, Ar-H), 8.02–
8.08 (m, 4H, Ar-H), 8.15 (s, 1H, C(8 or 2)H), 8.78 (s, 1H,
C(2 or 8)H), 9.32 (br s, 1H, NH); 13C NMR (500 MHz,

D. Vin˜a et al. / Tetrahedron 63 (2007) 2634–2646
2639
CDCl3) dC 38.59 (C-20), 73.43 (C-40), 74.34 (C-30), 85.15
(C-10), 123.88 (C-5), 127.88 (Ar-C), 128.45 (Ar-C), 128.73
(Ar-C), 129.64 (Ar-C), 132.69 (Ar-C), 133.40 (Ar-C),
141.80 (C-8), 149.69 (C-6), 151.45 (C-4), 152.52 (C-2),
164.74 (Bz-CO), 166.00 (Bz-CO); Exact mass calcd for
C23H19N5O4Na [M+Na]+ 452.1335, found 452.1339.
in 5 mL THF, which was cooled using dry-ice and acetone,
was added sodium hydride 80% (11.4 mg, 0.46 mmol).
The mixture was stirred for 10 min and the solution of
the triflate of the phosphonate (136 mg, 0.46 mmol) in
THF was slowly dropped into the reaction flask. Then the
mixture was slowly warmed up to room temperature. The re-
action was quenched with satd NaHCO3 and concentrated.
The residue was partitioned between H2O and CH2Cl2. The
organic layer was washed with water and brine, and concen-
trated in vacuo. The residue was purified by chromatography
on a silica gel column (CH2Cl2/MeOH¼98:2) to afford 11a
(25 mg, 0.06 mmol) as a colorless oil in 52% yield.
1
Compound 9b: H NMR (500 MHz, CDCl3) dH 2.90–2.96
(m, 1H, C(20)Hb), 3.05 (d, J¼15.4 Hz, 1H, C(20)Ha), 4.40
(dd, J1¼4.4 Hz, J2¼10.0 Hz, 1H, C(40)Hb), 4.46 (d, J¼
10.0 Hz, 1H, C(40)Ha), 5.74 (m, 1H, C(30)H), 6.56 (d,
J¼7.1 Hz, 1H, C(10)H), 7.38–7.61 (m, 6H, Ar-H), 7.74 (d,
J¼8.3 Hz, 2H, Ar-H), 8.03 (d, J¼7.6 Hz, 2H, Ar-H), 8.35
(s, 1H, C(8 or 2)H), 8.74 (s, 1H, C(2 or 8)H), 9.25 (br s,
1H, NH); 13C NMR (500 MHz, CDCl3) dC 38.63 (C-20),
73.45 (C-30), 74.80 (C-40), 85.19 (C-10), 123.44 (C-5),
127.90 (Ar-C), 128.59 (Ar-C), 128.76 (Ar-C), 129.36 (Ar-
C), 132.69 (Ar-C), 133.54 (Ar-C), 140.92 (C-8), 149.49
(C-6), 151.43 (C-4), 152.52 (C-2), 164.67 (Bz-CO), 165.79
(Bz-CO); Exact mass calcd for C23H19N5O4Na [M+Na]+
452.1335, found 452.1334.
Compound 11b was prepared as described for 11a using 10b
(60 mg, 0.27 mmol) as starting material. Compound 11b
(30 mg, 0.07 mmol) was obtained as a colorless oil in 52%
yield.
Compound 11a: 1H NMR (200 MHz, CDCl3) dH 1.35–1.38
(m, 12H, CH3), 2.62–2.74 (m, 1H, C(20)Ha), 2.93–3.02 (m,
1H, C(20)Hb), 3.78 (d, J¼8.6 Hz, 2H, PCH2), 4.17 (d,
J¼10 Hz, C(40)Hb), 4.37 (dd, 1H, J1¼4.2 Hz, J2¼9.8 Hz,
C(40)Ha), 4.64 (br s, 1H, C(30)H), 4.75–4.85 (m, 2H,
OCH(CH3)2), 5.91 (br s, 2H, NH2), 6.32 (t, 1H, J¼6.6 Hz,
C(10)H), 7.89 (s, 1H, C(8 or 2)H), 8.33 (s, 1H, C(2 or
8)H); 13C NMR (200 MHz, CDCl3) dC 23.97 (CH(CH3)3),
37.63 (C-20), 62.18 (PCH2), 65.55 (CH(CH3)3), 71.32
(CH(CH3)3), 73.35 (C-40), 81.73 (C-30), 85.68 (C-10),
139.49 (A-C(8)), 153.03 (A-C(2)), 155.61 (A-C(4)), C-5
and C-6 were obscured in noise; Exact mass calcd for
C16H26N5O5P1Na [M+Na]+ 422.1569, found 422.1572.
Elem. and calcd for C16H16N5O5P (MW, 399.1671) C:
48.10, H: 6.56, N: 17.54. Found C: 47.91, H: 6.24, N: 17.48.
4.1.5. 1-b-(Adenin-9-yl)-3-hydroxy-D-threo-tetrafura-
nose (10a) and 1-a-(adenin-9-yl)-3-hydroxy-D-erythro-
tetrafuranose (10b). A solution of 9a (90 mg, 0.21 mmol)
in methanol satd with ammonium (26 mL) was stirred at
room temperature overnight. The solution was concentrated
under vacuum and the residue was purified by column chro-
matography (CH2Cl2/MeOH¼9:1) to give compound 10a
(40 mg, 0.18 mmol) as a white solid in 86% yield.
Compound 10b was prepared as described for 10a using 9b
(110 mg, 0.26 mmol) as starting material. Compound 10b
(48 mg, 0.22 mmol) was obtained as a white solid in 86%
yield.
Compound 11b: 1H NMR (200 MHz, CDCl3) dH 1.25–1.35
(m, 12H, CH3), 2.57–2.62 (m, 2H, C(20)H2), 3.70 (dd,
J1¼1.4 Hz, J2¼8.8 Hz, 2H, PCH2), 4.04 (dd, J1¼4.2 Hz,
J2¼10.4 Hz, C(40)Ha), 4.37 (d, J¼10.4 Hz, C(40)Hb), 4.47
(br s, 1H, C(30)H), 4.71–4.78 (m, 2H, OCH(CH3)2), 5.75
(m, 2H, NH2), 6.47 (dd, J1¼2.7 Hz, J2¼7.0 Hz, C(10)H),
8.29 (s, 1H, C(8 or 2)), 8.34 (s, 1H, C(2 or 8)); 13C NMR
(200 MHz, CDCl3) dC 23.87 (CH(CH3)3), 38.56 (C-20),
62.41 (PCH2), 67.95 (CH(CH3)3), 71.46 (CH(CH3)3),
73.80 (C-40), 80.56 (C-30), 83.63 (C-10, 140.02 (A-C(8)),
153.03 (A-C(6)), 155.61 (A-C(2)); Exact mass calcd for
C16H26N5O5P1Na1 [M+Na]+ 422.1569, found 422.1573.
Elem. and calcd for C16H16N5O5P (MW, 399.1671) C:
48.10, H: 6.56, N: 17.54. Found C: 48.03, H: 6.32, N: 17.23.
1
Compound 10a: H NMR (200 MHz, DMSO-d6) dH 2.27–
2.34 (m, 1H, C(20)Ha), 2.78–2.85 (m, 1H, C(20)Hb), 3.76
(d, J¼8.8 Hz, 1H, C(40)Ha), 4.21 (dd, J1¼3.6 Hz, J2¼
8.8 Hz, 1H, C(40)Hb), 4.62 (br s, 1H, C(30)H), 5.20 (d,
J¼3.6 Hz, 1H, OH), 6.37 (t, J¼8.6 Hz, 1H, C(10)H), 7.26
(br s, 2H, NH2), 8.14 (s, 1H, C(8 or 2)), 8.30 (s, 1H, C(2
13
or 8)H); C NMR (200 MHz, DMSO-d6) dC 38.38 (C-20),
70.56 (C-30), 76.08 (C-40), 84.22 (C-10), 139.97 (A-C(8)),
152.78 (A-C(2)), 156.30 (A-C(6)), C-5 and C-4 were ob-
scured in noise; Exact mass calcd for C9H11N5O2Na
[M+Na]+ 244.0810, found 244.1191.
1
Compound 10b: H NMR (200 MHz, DMSO-d6) dH 2.23–
2.30 (m, 1H, C(20)Hb), 2.63–2.71 (m, 1H, C(20)Ha), 3.90–
3.93 (m, 2H, C(40)H2), 4.45 (br s, 1H, C(30)H), 5.80 (d,
J¼4.4 Hz, 1H, OH), 6.26 (dd, J1¼2.2 Hz, J2¼8.0 Hz, 1H,
C(10)H), 7.28 (s, 2H, NH2), 8.13 (s, 1H, C(8 or 2)H), 8.35
(s, 1H, C(2 or 8)H); 13C NMR (200 MHz, DMSO-d6) dC
38.49 (C-20), 66.99 (C-30), 74.04 (C-40), 80.90 (C-10),
118.01 (A-C(5)), 140.02 (A-C(8)), 152.52 (A-C(2)),
155.76 (A-C(6)), 160.97 (A-C(4)); Exact mass calcd for
C9H11N5O2Na [M+Na]+ 244.0810, found 244.1192.
4.1.7. 1-b-(Adenin-9-yl)-3-O-(phosphonomethyl)-D-
threo-tetrafuranose sodium salt (1a) and 1-a-(adenin-9-
yl)-3-O-(phosphonomethyl)-D-erythro-tetrafuranose
sodium salt (1b). To the solution of 11a (90 mg, 0.23 mmol)
and Et3N (0.33 mL) in DCM (9 mL) was added iodotri-
methylsilane (0.26 mL, 1.84 mmol) at 0 ꢀC. The reaction
mixture was stirred for 2 h. Then, the reaction was quenched
with 1.0 M TEAB solution. The mixture was concentrated
and the residue was purified by chromatography column
(CH2Cl2/MeOH¼1:2) to give the crude title compound. Pu-
rification using Sephadex-DEAE A-25 with gradient TEAB
solution from 0.01 to 0.8 M and ion exchange by Dowex-
Na+ resin afforded 1a as a white solid (37 mg, 0.11 mmol)
in 45% yield.
4.1.6. 1-b-(Adenin-9-yl)-3-O-(diisopropylphosphono-
methyl)-D-threo-tetrafuranose (11a) and 1-a-(adenin-9-
yl)-3-O-(diisopropylphosphonomethyl)-D-erythro-tetra-
furanose (11b). To the solution of 10a (50 mg, 0.23 mmol)

2640
D. Vin˜a et al. / Tetrahedron 63 (2007) 2634–2646
Compound 1b was prepared as described for 1a using 11b
(50 mg, 0.12 mmol) as starting material. Compound 1b
was obtained as a white solid (20 mg, 0.06 mmol) in 44%
yield.
isomer), 74.83 (C-30, 10b isomer), 75.38 (C-40, 10b isomer),
78.99 (C-20, 10a isomer), 83.21 (C-20, 10b isomer), 94.65
(C-10, 10a isomer), 99.90 (C-10, 10b isomer), 128.56 (Ar-
C), 129.22 (Ar-C), 129.71 (Ar-C), 133.47 (Ar-C), 165.55
(Bz-CO), 169.70 (CH3CO); Exact mass calcd for C19H28O6-
SiNa [M+Na]+ 403.1553, found 403.1554.
Compound 1a: 1H NMR (500 MHz, D2O) dH 2.81 (br s, 2H,
C(20)H2), 3.60–3.62 (m, 2H, PCH2), 4.20–4.25 (m, 2H,
C(40)H2), 4.64 (br s, 1H, C(30)H), 4.47 (t, J¼6.0 Hz, 1H,
C(10)H), 8.20 (s, 1H, A-C(8 or 2)H), 8.26 (s, 1H, C(2 or
8)H); 13C NMR (500 MHz, D2O) dC 39.40 (C-20), 68.76
(d, J¼61.1 Hz, PCH2), 75.76 (C-40), 83.39 (d, JP,C¼11.7 Hz,
C-30), 87.42 (C-10), 121.73 (A-C(5)), 142.99 (A-C(8)),
151.31 (A-C(6)), 155.35 (A-C(2)), 158.24 (A-C(4)); 31P
NMR (500 MHz, D2O) dP 14.74; Exact mass calcd for
C10H15N5O5P [M+H]+ 316.0811, found 316.0805.
4.1.9. 1-a-(Adenin-9-yl)-2-O-benzoyl-3-O-tertbutyldi-
methylsilyl-L-threose (16). To a mixture of 15 (1.0 g,
2.6 mmol) and N6-benzoyladenine (0.7 g, 5.2 mmol) in dry
MeCN (98 mL) was dropwise added SnCl4 (0.98 mL,
7.8 mmol) under N2 at 0 ꢀC. The reaction mixture was
stirred at room temperature for 1.5 h.
Then the reaction was quenched with satd NaHCO3 and
concentrated. The residue was partitioned between H2O
(50 mL) and EtOAc (200 mL). The organic layer was
washed with water and brine, and concentrated in vacuo.
The residue was purified by chromatography on a silica
gel column (CH2Cl2/MeOH¼98:2) to afford 16 (487 mg,
1.02 mmol) as a white solid in 39% yield.
Compound 1b: 1H NMR (500 MHz, D2O) dH 2.67 (d,
J¼14.9 Hz, 1H, C(20)Ha), 2.79 (m, 1H, C(20)Hb), 3.49–
3.57 (m, 2H, PH2), 4.09 (dd, J1¼10.2 Hz, J2¼4.1 Hz, 1H,
C(40)Ha), 4.34 (d, J¼10.2 Hz, 1H, C(40)Hb), 4.53–4.55 (m,
1H, C(30)H), 6.40 (dd, J1¼8.0 Hz, J2¼2.2 Hz, 1H,
C(10)H), 8.22 (s, 1H, C(2 or 8)H), 8.58 (s, 1H, C(8 or
2)H); 13C NMR (500 MHz, D2O) dC 39.73 (C-20), 69.54
(d, J¼61.5 Hz, PCH2), 76.49 (C-40), 82.27 (d, JP,C¼11.7 Hz,
C-30), 86.17 (C-10), 121.73 (A-C(5)), 144.17 (A-C(8)),
151.49 (A-C(6)), 155.34 (A-C(2)), 158.30 (A-C(4)); 31P
NMR (500 MHz, D2O) dP 14.02, Exact mass calcd for
C10H15N5O5P [M+H]+ 316.0811, found 316.0810.
1H NMR (200 MHz, CDCl3) dH 0.14 (d, J¼13.2 Hz, 6H,
SiCH3), 0.91 (s, 9H, CH3), 4.33–4.36 (m, 2H, C(40)H2),
4.53–4.55 (m, 1H, C(30)H), 5.62 (s, 1H, C(20)H), 5.92 (br
s, 2H, NH2), 6.48 (s, 1H, C(10)H), 7.47–7.65 (m, 3H, Ar-
H), 8.08–8.12 (m, 2H, Ar-H), 8.13 (s, 1H, C(2 or 8)H),
8.38 (s, 1H, C(8 or 2)H); 13C NMR (200 MHz, CDCl3) dC
ꢁ5.27 (SiCH3), ꢁ4.84 (SiCH3), 17.95 (C(CH3)3), 25.63
(C(CH3)3), 75.31 (C-40), 76.68 (C-30), 82.87 (C-20), 87.99
(C-10), 119.81 (A-C(5)), 128.73 (Ar-C), 128.75 (Ar-C),
130.07 (Ar-C), 133.95 (Ar-C), 139.69 (A-C(8)), 149.83
(A-C(4)), 153.32 (A-C(2)), 155.59 (A-C(4)), 165.24 (OB-
z(CO)); Exact mass calcd for C22H29N5O4SiNa [M+Na]+
478.1887, found 478.1911.
4.1.8. 1-O-Acetyl-2-O-benzoyl-3-O-tertbutyldimethyl-
silyl-L-threose (15). To the solution of 14 (1.0 g, 2.97 mmol)
in 10 mLTHF was slowly dropwise added 1.0 M diisopropyl
aluminum hydride (6 mL, 6 mmol) in toluene at ꢁ78 ꢀC,
and as soon as the starting material was completely con-
sumed (4 h) methanol (8 mL) was added over a period of
5 min to quench the reaction. The cooling bath was removed,
50 mL of satd aq sodium potassium tartrate solution and
100 mL of EtOAc were added, and the mixture stirred vigor-
ously for 3 h. The organic layer was washed with water and
brine, and concentrated in vacuo. The residue (950 mg,
2.80 mmol) was dissolved in 25 mL of Et3N and acetic an-
hydride was dropwise added (1.3 mL) followed by DMAP
(0.1 equiv, 34 mg) at 0 ꢀC. The reaction mixture was
warmed to room temperature for 3 h and concentrated in va-
cuo. The residue was purified by chromatography on a silica
gel column (n-hexane/EtOAc¼95:5, 9:1) to afford a mixture
of a/b isomers of 15 (880 mg, 2.3 mmol) in 78% global yield
for the two reactions. NMR showed that there were 66% of
a isomer and 34% of b isomer in the mixture of 15.
4.1.10. 1-a-(Adenin-9-yl)-3-O-tertbutyldimethylsilyl-
L-threose (17). A solution of 16 (480 mg, 1.05 mmol) in
MeOH satd with ammonia (79 mL) was stirred at room tem-
perature overnight. The mixture was concentrated and the
residue was purified by chromatography column (CH2Cl2/
MeOH¼92:8) to give compound 17 (370 mg, 1.04 mmol)
in 99% yield.
1H NMR (200 MHz, CDCl3) dH 0.10 (s, 6H, SiCH3), 0.82 (s,
9H, CH3), 2.47 (br s, 1H, OH), 4.20–4.23 (m, 1H, C(40)Ha),
4.39–4.46 (m, 3H, C(40)Hb, C(20)H, C(30)H), 1.82 (br s, 2H,
NH2), 6.10 (d, J¼2.4 Hz, 1H, C(10)H), 8.14 (s, 1H, C(2 or
8)H), 8.35 (s, 1H, C(8 or 2)H); 13C NMR (200 MHz,
CDCl3) dC ꢁ5.39 (SiCH3), 17.41 (C(CH3)3), 25.24
(C(CH3)3), 76.11 (C-40), 76.32 (C-30), 81.36 (C-20), 91.62
(C-10), 119.38 (A-C(5)), 138.96 (A-C(8)), 148.62 (A-
C(6)), 152.32 (A-C(2)), 155.26 (A-C(4)); Exact mass calcd
for C15H25N5O3SiNa [M+Na]+ 374.1624, found 374.1624.
1H NMR (200 MHz, CDCl3) dH 0.12 (d, J¼2.2 Hz, 6H,
SiCH3 (a isomer)), 0.17 (s, 6H, SiCH3 (b isomer)), 0.92 (t,
J¼3.0 Hz, 18 H, C-CH3, a+b isomer), 2.00 (s, 3H, CH3,
a isomer), 2.14 (s, 3H, CH3, b isomer), 3.80 (dd, J1¼4.4 Hz,
J2¼9.2 Hz, 1H, C(40)Ha, a isomer), 4.01 (dd, J1¼4.0 Hz,
J2¼9.2 Hz, 1H, C(40)Ha, b isomer), 4.26 (m, 2H, C(40)Hb,
a+b isomer), 4.46 (m, 1H, C(30)H, b isomer), 4.68 (m, 1H,
C(30)H, a isomer), 5.27 (m, 2H, C(20)H, a+b isomer), 6.22
(s, 1H, C(10)H, b isomer), 6.48 (d, J¼4.0 Hz, C(10)H, a iso-
mer), 7.26–7.60 (m, 6H, Ar-H), 8.00–8.06 (m, 4H, Ar-H);
13C NMR (200 MHz, CDCl3) dC ꢁ4.87 (SiCH3), ꢁ5.08
(SiCH3), 17.86 (C(CH3)3), 20.84, 21.00 (CO-CH3), 25.54
(C(CH3)3), 72.53 (C-30, 10a isomer), 73.38 (C-40, 10a
4.1.11. 1-a-(Adenin-9-yl)-2-azido-2-deoxy-3-O-tertbutyl-
dimethylsilyl-L-erythrose (18). Compound 17 (150 mg,
0.43 mmol) was dissolved in anhydrous DCM (2 mL) and
DMAP (130 mg) was added. The solution was cooled to
0 ꢀC and CF3SO2Cl (90 mL, 0.86 mmol) was added drop-
wise. After being stirred at 0 ꢀC for 2 h, the suspension
was taken up in AcOH/H2O(1:99)/CH2Cl2 and extracted
twice. The organic layers were washed with satd NaHCO3

D. Vin˜a et al. / Tetrahedron 63 (2007) 2634–2646
2641
solution and brine, dried over Na2SO4, evaporated and dried
under vacuum resulting in a white solid. The crude reaction
mixture was taken up in DMF (4 mL) and added to solid lith-
ium azide (90 mg, 2.15 mmol) under N2. The solution was
stirred at room temperature overnight, concentrated and
redissolved in CH2Cl2, and washed with NaHCO3 and brine.
The organic layer was dried over Na2SO4 and evaporated.
J1¼4.4 Hz, J2¼8 Hz, 1H, C(30)H), 4.74–4.85 (m, 2H,
OCH(CH3)2), 5.87 (br s, 2H, NH2), 6.47 (d, J¼6.4 Hz, 1H,
C(10)H), 8.23 (s, 1H, C(2 or 8)H), 8.35 (s, 1H, C(8 or
2)H); 13C NMR (200 MHz, CDCl3) dC 24.05 (CH(CH3)3),
63.14 (C-20), 65.70 (PCH2), 70.97 (C-40), 71.64 (CH(CH3)3),
80.25 (d, JP,C¼30.4 Hz, C-30), 82.81 (C-10), 118.79 (A-
C(5)), 140.54 (A-C(8)), 150.19 (A-C(6)), 153.29 (A-C(2)),
155.56 (A-C(4)); Exact mass calcd for C16H25N8O5PNa
[M+Na]+ 463.1583, found 463.1621. Elem. and calcd for
C10H25N8O5P (MW, 440.1685) C: 43.62, H: 5.72, N:
25.45. Found C: 43.93, H: 5.41, N: 25.07.
Purification by preparative TLC using as eluent CH2Cl2/
MeOH¼9:1 afforded 18 as a white solid (60 mg, 0.16 mmol)
in 37% yield.
1H NMR (200 MHz, CDCl3) dH 0.19 (d, J¼1.8 Hz, 6H,
SiCH3), 0.96 (s, 9H, CH3), 4.15 (d, J¼5.2 Hz, 2H,
C(40)H2), 4.29 (m, 1H, C(20)H), 4.72 (m, 1H, C(30)H), 5.71
(br s, 2H, NH2), 6.48 (d, J¼5.4 Hz, 1H, C(10)H), 8.22 (s,
1H, C(2 or 8)H), 8.38 (s, 1H, C(8 or 2)H); 13C NMR
(200 MHz, CDCl3) dC ꢁ6.52 (SiCH3), 18.45 (C(CH3)3),
24.23 (C(CH3)3), 62.56 (C-20), 71.06 (C-40), 71.79 (C-30),
81.38 (C-10), 120.25 (C(5)), 139.08 (A-C(8)), 146.40
(C(6)), 151.74 (C(2)), 153.95 (C(4)); Exact mass calcd for
C15H24N8O2SiNa [M+Na]+ 399.1869, found 399.1703.
4.1.14. 1-a-(Adenin-9-yl)-2-azido-2-deoxy-3-O-(phos-
phonomethyl)-L-erythrose sodium salt (2). To a solution
of 20 (80 mg, 0.18 mmol) and Et3N (0.26 mL) in DCM
(7.2 mL) was added iodotrimethylsilane (0.20 mL) at 0 ꢀC.
The reaction mixture was stirred for 4 h at room temperature.
The reaction was quenched with 1.0 M TEAB solution. The
mixture was concentrated and the residue was purified by
column chromatography (CH2Cl2/MeOH¼8:2, 5:5, 2:8) to
give crude title compound. Purification using Sephadex-
DEAE A-25 with gradient TEAB solution from 0.01 to
1.0 M and ion exchanges by Dowex-Na+ resin afforded 2
(30 mg, 0.07 mmol) as a white solid in 41% yield.
4.1.12. 1-a-(Adenin-9-yl)-2-azido-2-deoxy-L-erythrose
(19). To the solution of 18 (120 mg, 0.32 mmol) in 3 mL
of THF, was added at room temperature TBAF (64 mL,
0.64 mmol). The solution was stirred at room temperature
for 2 h. The mixture was concentrated and the residue was
purified by chromatography on a silica gel column
(CH2Cl2/MeOH¼9:1) to afford 19 (70 mg, 0.27 mmol) as
a white solid in 84% yield.
1H NMR (500 MHz, D2O) dH 3.70 (d, J¼10.8 Hz, 2H, CH2-
P), 4.19 (dd, J1¼10.2 Hz, J2¼4.2 Hz, 1H, C(40)Hb), 4.52
(dd, J1¼10.2 Hz, J2¼2.3 Hz, 1H, C(40)Ha), 4.59 (m, 1H,
C(30)H), 4.78 (dd, J1¼5.1 Hz, J2¼6.3 Hz, 1H, C(20)H),
6.47 (d, J¼6.3 Hz, 1H, C(10)H), 8.24 (s, 1H, A-C(8)H), 8.53
(s, 1H, A-C(2)H); 13C NMR (500 MHz, D2O) dC 66.02 (C-
20), 70.77 (d, PCH2), 74.24 (C-40), 81.86 (C-30), 85.92 (C-
10), 120.29 (A-C(5)), 144.72 (A-C(8)), 151.55 (A-C(6)),
155.54 (A-C(2)), 158.29 (A-C(4)); 31P NMR (500 MHz,
D2O) dP 13.46; Exact mass calcd for C10H14N8O5P
[M+H]+ 357.0825, found 357.0829.
1H NMR (200 MHz, DMSO-d6) dH 4.04–4.06 (m, 2H,
C(40)H2), 4.54–4.63 (m, 2H, C(20)H, C(30)H), 6.31 (d,
J¼4.6 Hz, 1H, OH), 6.37 (d, J¼6.2 Hz, 1H, C(10)H), 7.32
(br s, 2H, NH2), 8.16 (s, 1H, A-C(2 or 8)H), 8.29 (s, 1H,
A-C(8 or 2)H); 13C NMR (200 MHz, DMSO-d6) dC 63.63
(C-20), 70.52 (C-40), 74.25 (C-30), 82.84 (C-10), 119.02 (A-
C(5)), 140.97 (A-C(8)), 150.89 (A-C(6)), 153.35 (A-C(2)),
156.75 (A-C(4)); Exact mass calcd for C9H10N8O2Na1
[M+Na]+ 285.0825, found 285.0839. Elem. and calcd for
C9H10N8O2 (MW, 262.0927) C: 41.21, H: 3.85, N: 42.74.
Found C: 41.54, H: 3.71, N: 42.48.
4.1.15. 1,2:5,6-Di-O-isopropylidene-3-O-(diisopropyl-
phosphonomethyl)-a-D-galactofuranose (22). To a solu-
tion of 1,2:5,6-di-O-isopropylidene-a-D-galactofuranose 21
(1.16 g, 4.46 mmol) in dried THF (25 mL) was added so-
dium hydride (80% dispersion in mineral oil, 268 mg,
8.92 mmol) at ꢁ78 ꢀC. Then the solution of the triflate of
diisopropylphosphonomethanol3 (2.64 g, 8.92 mmol) in
dried THF (5 mL) was dropwise added, and the reaction
mixture was slowly warmed to room temperature. The reac-
tion was quenched with satd NaHCO3 and concentrated. The
residue was partitioned between H2O and EtOAc. The or-
ganic layer was washed with water and brine, dried over
Na2SO4, and concentrated in vacuo. The residue was purified
by chromatography on a silica gel column (DCM/
MeOH¼60:1) to afford 22 (1.64 g, 3.75 mmol, 84%) as
colorless oil.
4.1.13. 1-a-(Adenin-9-yl)-2-azido-2-deoxy-3-O-(diisopro-
pylphosphonomethyl)-L-erythrose (20). To the solution of
19 (30 mg, 0.11 mmol) in 3 mL of THF, which was cooled
using dry-ice in acetone, was added sodium hydride 80%
(7 mg, 0.22 mmol). The mixture was stirred for 10 min
and the solution of the trifluoromethanesulfonate of the
phosphonate reagent (65 mg, 0.22 mmol) in THF was slowly
dropped into the reaction flask. Then the mixture was slowly
warmed up to room temperature. The reaction was quenched
with satd NaHCO3 and concentrated. The residue was parti-
tioned between H2O and CH2Cl2. The organic layer was
washed with water and brine, and concentrated in vacuo.
The residue was purified by chromatography on a silica
gel column (CH2Cl2/MeOH¼94:6) to afford 20 (37 mg,
0.08 mmol) in 74% yield.
1H NMR (200 MHz, CDCl3) dH 1.29–1.51 (m, 24H, CH3),
3.67–3.89 (m, 5H, PCH2, C(40)H, C(60)H2), 4.42 (dd,
J1¼4.2 Hz, J2¼6.6 Hz, C(30)H), 4.58 (d, J¼4.0 Hz, C(20)H),
4.22–4.31 (m, 1H, C(50)H), 4.64–4.81 (m, 2H, CH(CH3)2),
5.79 (d, J¼4.0 Hz, C(10)H); 13C NMR (200 MHz, CDCl3)
dC 23.91 (CH3), 25.22 (CH3), 26.71 (CH3), 27.38 (CH3),
64.89 (d, JP,C¼170.9 Hz, PCH2), 65.59 (C-60), 71.17
(CH(CH3)2), 71.29 (CH(CH3)2), 75.48 (C-50), 83.44 (C-20),
84.86 (C-40), 84.52 (d, JP,C¼12.2 Hz), 104.92 (C-10), 109.8
1H NMR (200 MHz, CDCl3) dH 1.33–1.39 (m, 12H, CH3),
3.80–3.99 (m, 2H, PCH2), 4.16 (dd, J1¼4.8 Hz, J2¼10.2 Hz,
1H, C(40)Ha), 4.32–4.51 (m, 2H, C(20)H, C(40)Hb), 4.57 (dd,

2642
D. Vin˜a et al. / Tetrahedron 63 (2007) 2634–2646
(C(O)2), 113.88 (C(O)2); Exact mass (EI) calcd for
C16H29O8P [MꢁC3H6O] 380.1600, found 380.1598.
To the solution of 3-O-(diisopropylphosphonomethyl)-5-
deoxy-5-chloromethylene-a,b-L-arabinofuranose (450 mg,
1.25 mmol) in 100 mL pyridine was added dropwise BzCl
(0.31 mL, 2.75 mmol) at 0 ꢀC. The reaction mixture was
warmed to room temperature and stirred overnight. The
reaction mixture was concentrated and coevaporated with
20 mL of toluene two times in vacuo. The residue was parti-
tioned between H2O (30 mL) and EtOAc (200 mL). The
organic layer was washed with water and brine, dried over
Na2SO4, and concentrated in vacuo. The residue was purified
by chromatography on a silica gel column (n-hexane/
EtOAc¼2:1) to afford 25 (610 mg, 1.07 mmol) as colorless
oil in 86% yield.
4.1.16. 1,2-O-Isopropylidene-3-O-(diisopropylphospho-
nomethyl)-5-oxo-b-L-arabinofuranose (23). To a solution
of 22 (5.00 g, 10.3 mmol) in 5 mL of ethyl acetate was added
a well-stirred suspension of NaIO4 (2.21 g, 10.3 mmol) and
H5IO6 (1.17 g, 5.16 mmol) in 100 mL of ethyl acetate. After
stirring for 5 h at room temperature, the mixture was filtered.
The filtrate was concentrated to dry, affording 23 (3.74 g,
10.2 mmol) as colorless oil in 99% yield.
1H NMR (200 MHz, CDCl3) dH 1.28–1.42 (m, 18H, CH3),
3.77 (s, 1H, PCHa), 3.82 (s, 1H, PCHb), 4.28 (s, 1H,
C(40)H), 4.50 (s, 1H, C(30)H), 4.63 (d, J¼3.5 Hz, 1H,
C(20)H), 4.60–4.90 (m, 2H, CH(CH3)2), 6.02 (d, J¼3.5 Hz,
1H, C(10)H), 9.75 (s, 1H, C(50)H); 13C NMR (200 MHz,
CDCl3) dC 22.41 (CH3), 25.66 (CH3), 26.21 (CH3), 64.55
(d, JP,C¼169.4 Hz, PCH2), 71.62 (CH(CH3)2), 71.29
(CH(CH3)2), 82.51 (C-20), 87.50 (d, JP,C¼10.7 Hz, (C-30)),
88.58 (C-40), 106.7 (C-10), 112.67 (C(O)2), 201.49 (C-50).
1H NMR (200 MHz, CDCl3) dH 1.30–1.34 (m, 12H, CH3),
3.89–4.21 (m, 3H, PCH2, C(40)H), 4.54–4.86 (m, 3H,
C(30)H, CH(CH3)2), 5.56–5.62 (m, 1H, C(20)H), 5.94–6.29
(m, 1H, C(50)H), 6.39–6.51 (m, 1H, C(60)H, (10)H), 6.61–
6.75 (m, 1H, C(60)H), 7.35–7.68 (m, 9H, Ar-H), 7.92–8.18
(m, 6H, Ar-H); 13C NMR (200 MHz, CDCl3) dC 23.95
(CH3), 65.41 (d, JP,C¼167.9 Hz, PCH2), 71.42 (CH(CH3)2),
80.37, 80.55 (C-40), 83.28 (C-20), 89.03 (d, JP,C¼10.7 Hz,
(C-30)), 94.60, 99.82 (C-10), 122.89 (C-50), 128.51, 128.69,
129.78, 129.90, 130.15 (Ar-C), 131.63 (C-60), 133.67,
133.88 (Ar-C), 165.14 (BzCO); Exact mass calcd for
C27H32Cl1O9PNa [M+Na]+ 589.1370, found 589.1375.
4.1.17. 1,2-O-Isopropylidene-3-O-(diisopropylphos-
phonomethyl)-5-deoxy-5-chloromethylene-b-L-arabino-
furanose (24). A suspension of (chloromethyl)triphenyl-
phosphonium chloride (3.82 g, 11 mmol) in anhydrous
THF (50 mL) was cooled to ꢁ78 ꢀC and treated with a solu-
tion of n-BuLi in hexane (1.6 M, 6.87 mL, 11 mmol). After
1 h of stirring, a solution of aldehyde 23 (1.04 g,
2.75 mmol) in dry THF (50 mL) was added. The temperature
of the bath was raised to 0 ꢀC and stirring continued for 3 h.
Following cautious addition of an aq satd solution of NH4Cl,
the reaction mixture was extracted with EtOAc (2ꢂ150 mL).
The combined organic layer was washed with brine, dried
over Na2SO4, and concentrated in vacuo. The residue was
purified by chromatography on a silica gel column (DCM/
MeOH¼98:2) to afford 24 contaminated with triphenylphos-
phine. A sample for NMR analysis was prepared by prepara-
tive TLC.
4.1.19. 1-(N6-Benzoyladenin-9-yl)-2-O-benzoyl-3-O-
(diisopropylphosphonomethyl)-5-deoxy-5-chloromethyl-
ene-a-L-arabinofuranose (26). To a mixture of 25 (270 mg,
0.48 mmol) and silylated N6-benzoyladenine (225 mg,
0.96 mmol) in dry MeCN (30 mL) was dropwise added
SnCl4 (0.11 mL, 0.94 mmol) under N2 at room temperature.
The reaction mixture was stirred at room temperature for
4–5 h. Then the reaction was quenched with satd NaHCO3
and concentrated. The residue was partitioned between
H2O (20 mL) and EtOAc (100 mL). The organic layer was
washed with water and brine, dried over Na2SO4, and con-
centrated in vacuo. The residue was purified by chromato-
graphy on a silica gel column (DCM/MeOH¼40:1) to
afford 26 (239 mg, 0.35 mmol) as a colorless amorphous
solid in 73% yield.
1H NMR (200 MHz, CDCl3) dH 1.32–1.52 (m, 15H, CH3),
1.72 (s, 3H, CH3), 3.62–4.04 (m, 3H, PCH2, C(40)H),
4.52–5.12 (m, 4H, C(30)H, C(20)H, CH(CH3)2), 5.93–5.95
(m, 1H, C(10)H), 6.04–6.16 (m, 1H, C(50)H), 6.31–6.38
(m, 1H, C(60)H); 13C NMR (200 MHz, CDCl3) dC 23.94
(CH3), 26.16 (CH3), 26.65 (CH3), 64.70 (d, JP,C¼170.9 Hz,
PCH2), 71.32 (CH(CH3)2), 83.01 (C-20), 84.01 (C-40), 88.40
(d, JP,C¼10.5 Hz, C-30), 105.83 (C-10), 113.15 (C(O)2),
121.80 (C-50), 133.61 (C-60); Exact mass calcd for
C16H28Cl1O7PNa [M+Na]+ 421.1159, found 421.1165.
1H NMR (200 MHz, CDCl3) dH 1.30–1.37 (m, 12H, CH3),
3.89–4.14 (m, 2H, PCH2), 4.38–4.40 (m, 1H, C(40)H),
4.56–4.84 (m, 2H, CH(CH3)2), 4.85–4.92 (m, 0.8H,
C(30)H), 5.60–5.62 (m, 0.2H, C(30)H), 5.94–6.75 (m, 4H,
C(20)H, C(10)H, C(50)H, C(60)H), 7.40–7.70 (m, 6H, Ar-
H), 8.03–8.07 (m, 4H, Ar-H), 8.42 (s, 0.8H, C(8)H), 8.54
(s, 0.2H, C(8)H), 8.84 (s, 1H, C(2)H), 9.12 (s, 1H, NH);
13C NMR (200 MHz, CDCl3) dC 23.94 (CH3), 65.65 (d,
JP,C¼181.5 Hz, PCH2), 71.51, 71.63 (CH(CH3)2), 80.28
(C-40), 83.98 (C-20), 87.90 (C-30), 88.11 (C-10), 122.00 (C-
5), 123.20 (C-50), 127.90, 128.81, 128.93, 129.37, 129.99
(Ar-C), 132.85 (C-60), 134.18, (Ar-C), 141.95 (C-8),
149.72 (C-6), 153.09 (C-2), 165.32 (CO), C-4 was obscured
in noise peak; Exact mass calcd for C32H35Cl1N5O8PNa
[M+Na]+ 706.1809, found 706.1806.
4.1.18. 1,2-Di-O-benzoyl-3-O-(diisopropylphosphono-
methyl)-5-deoxy-5-chloromethylene-a,b-L-arabinofura-
nose (25). A solution of crude product 24 in TFA/H2O (3:1,
20 mL) was allowed to stand at room temperature for 2 h.
The reaction mixture was neutralized with satd NaHCO3
solution. The mixture was partitioned between the DCM
(400 mL) and water (20 mL). The organic layer was washed
with water and brine, dried over Na2SO4, and then concen-
trated in vacuo. The residue was purified by chromatography
on silica gel (DCM/MeOH¼20:1) to give the debenzoylated
compound (460 g, 1.28 mmol) as a colorless amorphous
solid in 46% of yield calculated from compound 23.
4.1.20. 1-(Adenin-9-yl)-3-O-(diisopropylphosphono-
methyl)-5-deoxy-5-chloromethylene-a-L-arabinofura-
nose (27). A solution of 26 (328 mg, 0.48 mmol) in MeOH
satd with ammonia (100 mL) was stirred at room

D. Vin˜a et al. / Tetrahedron 63 (2007) 2634–2646
2643
temperature overnight. The mixture was concentrated and
the residue was purified by column chromatography
(CH2Cl2/MeOH¼20:1) to give compound 27 (210 mg,
0.42 mmol) as a white powder in 88% yield.
C(40)H), 4.08–4.17 (m, 2H, C(50)H2), 4.64 (d, J¼4.0 Hz,
1H, C(20)H), 4.65–4.92 (m, 2H, CH(CH3)2), 5.87 (d,
J¼4.0 Hz, 1H, C(10)H); 13C NMR (200 MHz, CDCl3) dC
23.88 (CH3), 26.28 (CH3), 27.04 (CH3), 62.40 (C-50), 64.54
(d, JP,C¼169.4 Hz, PCH2), 71.32 (CH(CH3)2), 71.45
1H NMR (200 MHz, CDCl3) dH 1.25–1.31 (m, 12H, CH3),
3.84–4.08 (m, 3H, PCH2, C(40)H), 4.57–4.80 (m, 2H,
CH(CH3)2), 4.85–4.92 (m, 1H, C(30)H), 5.04 (dd, J1¼
5.1 Hz, J2¼4.7 Hz, 0.86H, C(20)H), 5.42–5.47 (m, 0.16H,
C(20)H), 5.99–6.43 (m, 3H, C(10)H, C(50)H, C(60)H), 6.59
(br s, 1.68H, NH), 6.45 (br s, 0.28H, NH), 7.97 (s, 0.86H,
C(8)H), 8.07 (s, 0.24H, C(8)H), 8.16 (s, 1H, C(2)H); 13C
(CH(CH3)2), 84.86 (C-20), 84.98 (C-40), 85.79 (d, JP,C
¼
10.6 Hz, C-30), 105.3 (C-10), 113.03 (C(O)2); Exact mass
calcd for C15H29O8PNa [M+Na]+ 391.1498, found 391.1492.
4.1.23. 1,2-O-Isopropylidene-3-O-(diisopropylphospho-
nomethyl)-5-O-benzoyl-b-L-arabinofuranose (29). To
a solution of 28 (3.20 g, 8.7 mmol), DMAP (106 mg,
0.87 mmol) and Et3N (3.7 mL, 26.1 mmol) in 50 mL of
DCM was added dropwise BzCl (1.3 mL, mmol) at 0 ꢀC.
The reaction mixture was warmed to room temperature
and stirred overnight. The reaction mixture was concentrated
in vacuo. The residue was partitioned between H2O (50 mL)
and EtOAc (250 mL). The organic layer was washed with
water and brine, dried over Na2SO4, and concentrated in va-
cuo. The residue was purified by chromatography on a silica
gel column (DCM/MeOH¼98:2) to afford 29 (2.81 g,
5.9 mmol) as colorless oil in 68% yield.
NMR (200 MHz, CDCl3) dC 23.82 (CH3), 65.59 (d, JP,C
¼
181.5 Hz, PCH2), 71.57, 71.72, 71.87, 71.99 (CH(CH3)2),
78.76 (C-40), 81.46 (C-20), 89.74 (C-30, C-10), 119.68 (C-
5), 122.98 (C-50), 129.78, 130.45 (C-60), 139.34 (C-8),
149.12 (C-6), 152.76 (C-2), 155.64 (C-4); Exact mass calcd
for C18H28Cl1N5O6P [M+H]+ 476.1466, found 476.1465.
4.1.21. 1-(Adenin-9-yl)-3-O-(phosphonomethyl)-5-deoxy-
5-chloromethylene-a-L-arabinofuranose disodium salt
(3). To a solution of 27 (140 mg, 0.30 mmol) and Et3N
(1 mL) in DCM (25 mL) was added iodotrimethylsilane
(0.43 mL, 3.0 mmol) at 0 ꢀC. The reaction mixture was
stirred for 2 h. The reaction was quenched with 1.0 M
TEAB solution. The mixture was concentrated and the resi-
due was purified by column chromatography (CH2Cl2/
MeOH¼2:1, 1:1, 1:2) to give crude 1-(adenin-9-yl)-3-O-
(phosphonomethyl)-5-deoxy-5-chloromethylene-a-L-arabi-
nofuranose triethylammonium salt. Purification using
Sephadex-DEAE A-25 with gradient TEAB solution from
0.01 to 0.5 M and ion exchanges by the Dowex-Na+ resin af-
forded 3 (25 mg, 0.06 mmol) as a colorless solid after pre-
cipitation from diethylether in 20% yield.
1H NMR (200 MHz, CDCl3) dH 1.31–1.36 (m, 15H, CH3),
1.56 (s, 1H, CH3), 3.80 (s, 1H, PCHa), 3.85 (s, 1H, PCHb),
4.11 (d, J¼2.6 Hz, 1H, C(40)H), 4.34–4.42 (m, 1H,
C(50)Ha), 4.50 (s, 1H, C(30)H), 4.53 (d, J¼1.5 Hz, 1H,
C(50)Hb), 4.68 (d, J¼3.7 Hz, 1H, C(20)H), 4.67–4.84 (m,
2H, CH(CH3)2), 5.92 (d, J¼3.7 Hz, 1H, C(10)H); 13C
NMR (200 MHz, CDCl3) dC 23.94 (CH3), 26.13 (CH3),
27.04 (CH3), 64.31 (C-50), 64.68 (d, JP,C¼170.9 Hz,
PCH2), 71.36 (CH(CH3)2), 82.19 (C-40), 84.16 (C-20),
85.80 (d, JP,C¼12.2 Hz, C-30), 105.80 (C-10), 113.24
(C(O)2), 128.45 (Ar-C), 128.78 (Ar-C), 133.24 (Ar-C),
166.23 (BzCO); Exact mass (EI) calcd for C19H27O8P
[MꢁC3H6O] 414.1444, found 414.1437.
1H NMR (200 MHz, D2O) dH 3.23–3.54 (m, 2H, PCH2),
3.76–3.87 (m, 1H, C(40)H), 4.55–4.61 (m, 1.84H, C(30)H,
C(20)H), 5.06–5.13 (m, 0.16H, C(20)H), 5.76–5.96 (m, 2H,
C(10)H, C(50)H), 6.13 (d, J¼9.3 Hz, 0.16H, C(60)H), 4.25
(d, J¼13.2 Hz, 0.84H, C(60)H), 7.77 (s, 1H, C(8)H), 8.01
(s, 0.84H, C(2)H), 8.04 (s, 0.84H, C(2)H); 13C NMR
(200 MHz, CDCl3) dC 70.62 (d, JP,C¼151.1 Hz, PCH2),
80.04, 81.29 (C-40), 84.20 (C-20), 90.06, 90.97 (C-10), 91.06
(d, JP,C¼9.2 Hz, C-30), 120.63 (C-5), 125.27, 125.61 (C-50),
130.86, 132.56 (C-60), 142.82 (C-8), 150.74 (C-6), 154.84
(C-2), 157.51 (C-4); Exact mass calcd for C12H16N5O6P
[M+H]+ 392.0527, found 392.0521.
4.1.24. 1,2,5-Tri-O-benzoyl-3-O-(diisopropylphosphono-
methyl)-a-L-arabinofuranose (30a) and 1,2,5-tri-O-ben-
zoyl-3-O-(diisopropylphosphonomethyl)-b-L-arabino-
furanose (30b). A solution of 29 (2.71 g, 5.7 mmol) in TFA/
H2O (3:1, 20 mL) was allowed to stand at room temperature
for 2 h. The reaction mixture was neutralized with satd
NaHCO3 solution. Then the mixture was partitioned be-
tween DCM (400 mL) and water (20 mL). The organic layer
was washed with water and brine, dried over Na2SO4, and
then concentrated in vacuo. The residue was purified by
chromatography on silica gel (DCM/MeOH¼30:1) to give
3-O-(diisopropylphosphonomethyl)-a,b-L-arabinofuranose
(2.21 g, 5.1 mmol) as colorless oil in 89% yield.
4.1.22. 1,2-O-Isopropylidene-3-O-(diisopropylphospho-
nomethyl)-b-L-arabinofuranose (28). To a solution of 23
(3.70 g, 10.1 mmol) in MeOH (50 mL) was added NaBH4
(1.15 g, 30.9 mmol) at room temperature. After the mixture
was stirred at room temperature for 2 h, AcOH (10%) was
added to neutralize the reaction mixture. Then the mixture
was concentrated and partitioned between EtOAc (200 mL)
and H2O (50 mL). The organic phase was washed with brine,
dried with Na2SO4, and concentrated to dryness. The residue
was purified by chromatography on a silica gel column
(DCM/MeOH¼95:1) to afford 28 (3.40 g, 10.0 mmol) as
colorless oil in 99% yield.
To the solution of 3-O-(diisopropylphosphonomethyl)-a,b-
L-arabinofuranose (2.11 g, 5.1 mmol) in 100 mL pyridine
was added dropwise BzCl (14.0 mL, 12.2 mmol) at 0 ꢀC.
The reaction mixture was warmed to room temperature
and stirred overnight. The reaction mixture was concentrated
and coevaporated with 20 mL toluene two times in vacuo.
The residue was partitioned between H2O (20 mL) and
EtOAc (150 mL). The organic layer was washed with water
and brine, dried over Na2SO4, and concentrated in vacuo.
The residue was purified by chromatography on a silica
gel column (DCM/MeOH¼95:5) to afford 30a and 30b
(2.64 g, 4.1 mmol) as colorless oil in 81% yield. The H-10
1H NMR (200 MHz, CDCl3) dH 1.33–1.36 (m, 15H, CH3),
1.53 (s, 1H, CH3), 3.71–3.94 (m, 4H, PCH2, C(30)H,

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D. Vin˜a et al. / Tetrahedron 63 (2007) 2634–2646
proton of 30a appeared as a singlet while the H-10 proton of
30b is doublet (J¼4.7 Hz).
(383 mg, 0.50 mmol) in MeOH satd with ammonia
(100 mL) was stirred at room temperature overnight. The
mixture was concentrated and the residue was purified by
column chromatography (CH2Cl2/MeOH¼20:1) to give
compound 32 (196 mg, 0.44 mmol) as a colorless amor-
phous solid in 87% yield.
Compound 30a: 1H NMR (200 MHz, CDCl3) dH 1.26–1.39
(m, 12H, CH3), 4.06–4.38 (m, 3H, PCH2, C(40)H), 4.57 (dd,
J1¼12.8 Hz, J2¼5.5 Hz, 1H, C(50)Ha), 4.68–4.91 (m, 4H,
C(30)H, C(50)Hb, CH(CH3)2), 5.62 (s, 1H, C(20)H), 6.72 (s,
1H, C(10)H), 7.12–7.65 (m, 9H, Ar-H), 7.96–8.16 (m, 6H,
Ar-H); 13C NMR (200 MHz, CDCl3) dC 23.79 (CH3),
63.22 (C-50), 65.24 (d, JP,C¼169.4 Hz, PCH2), 71.48
(CH(CH3)2), 71.60 (CH(CH3)2), 80.61 (C-40), 83.31 (C-20),
85.98 (d, JP,C¼12.2 Hz, C-30), 100.04 (C-10), 128.23–
129.36 (Ar-C), 129.63–130.06 (Ar-C), 133.06–133.67 (Ar-
C), 164.87, 165.32, 166.14 (BzCO); Exact mass (EI) calcd
for C33H37O11P [M] 640.2073, found 640.2072.
1H NMR (200 MHz, CDCl3) dH 1.27–1.32 (m, 12H, CH3),
3.80–3.98 (m, 4H, PCH2, C(50)H2), 4.30 (t, J¼5.1 Hz,
C(40)H), 4.60–4.80 (m, 2H, CH(CH3)2), 4.90 (dd,
J1¼4.4 Hz, J2¼4.5 Hz, 1H, C(20)H), 6.04 (d, J¼4.4 Hz,
1H, C(10)H), 6.32 (br s, 2H, NH), 8.04 (s, 1H, C(8)H),
8.16 (s, 1H, C(2)H); 13C NMR (200 MHz, CDCl3) dC
23.88 (CH3), 61.58 (C-50), 65.40 (d, JP,C¼167.8 Hz,
PCH2), 71.75 (m, CH(CH3)2), 79.27 (C-20), 83.98 (C-40),
85.73 (d, JP,C¼9.8 Hz, C-30), 89.96 (C-10), 119.43 (C-5),
139.25 (C-8), 149.12 (C-6), 152.73 (C-2), 155.49 (C-4);
Exact mass calcd for C17H29N5O7P [M+H]+ 446.1805,
found 446.1808.
Compound 30b: 1H NMR (200 MHz, CDCl3) dH 1.26–1.36
(m, 12H, CH3), 3.96–4.12 (m, 3H, PCH2, C(40)H), 4.48–4.82
(m, 4H, C(50)H2, C(30)H, CH(CH3)2), 5.63 (dd, J1¼6.9 Hz,
J2¼4.7 Hz, 1H, C(20)H), 6.77 (d, J¼4.7 Hz, 1H, C(10)H),
7.23–7.61 (m, 9H, Ar-H), 7.86–8.12 (m, 6H, Ar-H); 13C
NMR (200 MHz, CDCl3) dC 23.82 (CH3), 64.28 (C-50),
65.12 (d, JP,C¼169.3 Hz, PCH2), 71.38 (CH(CH3)2), 71.51
(CH(CH3)2), 77.52 (C-20), 79.46 (C-40), 82.52 (d,
JP,C¼12.1 Hz, C-30), 94.45 (C-10), 128.39, 128.51, 129.24,
129.69, 132.76, 133.15, 133.39, 133.63 (Ar-C), 164.81,
165.32, 166.23 (BzCO); Exact mass (EI) calcd for
C33H37O11P [M] 640.2073, found 640.2076.
4.1.27. 1-(Adenin-9-yl)-3-O-(phosphonomethyl)-a-L-ara-
binofuranose disodium salt (4). To a solution of 32
(60 mg, 0.13 mmol) and Et3N (1 mL) in DCM (25 mL)
was added iodotrimethylsilane (0.17 mL, 1.30 mmol) at
0 ꢀC. After stirring for 2 h, the reaction mixture was
quenched with 1.0 M TEAB solution. The mixture was con-
centrated and the residue was purified by column chromato-
graphy (CH2Cl2/MeOH¼2:1, 1:1, 1:2) to give crude 4 as
triethylammonium salt. Purification using Sephadex-DEAE
A-25 with gradient TEAB solution from 0.01 to 0.5 M and
ion exchanges by the Dowex-Na+ resin afforded 4 (20 mg,
0.05 mmol) as a colorless solid after precipitation from di-
ethylether in 38% yield.
4.1.25. 1-(N6-Benzoyladenin-9-yl)-2,5-di-O-benzoyl-3-O-
(diisopropylphosphonomethyl)-a-L-arabinofuranose
(31). To a mixture of 30a and 30b (720 mg, 1.12 mmol) and
N6-benzoyladenine (537 mg, 2.24 mmol) in dry MeCN
(50 mL) was added dropwise SnCl4 (522 mL, 4.48 mmol)
under N2 at room temperature. The reaction mixture was
stirred at room temperature for 24 h. Then the reaction
was quenched with satd NaHCO3 and concentrated. The resi-
due was partitioned between H2O (20 mL) and EtOAc
(100 mL). The organic layer was washed with water and
brine, dried over Na2SO4, and concentrated in vacuo. The
residue was purified by chromatography on a silica gel col-
umn (DCM/MeOH¼100:1, 40:1) to afford 31 (520 mg,
0.69 mmol) as a colorless amorphous solid in 61% yield.
1H NMR (500 MHz, CDCl3) dH 3.73 (dd, J1¼9.0 Hz,
J2¼12.9 Hz, 1H, PCHa), 3.76 (dd, J1¼9.0 Hz, J2¼12.9 Hz,
1H, PCHb), 3.86 (dd, J1¼12.7 Hz, J2¼5.1 Hz, 1H,
C(50)Ha), 3.89 (dd, J1¼12.7 Hz, J2¼3.9 Hz, 1H, C(50)Hb),
4.18 (dd, J1¼3.9 Hz, J2¼5.1 Hz, 1H, C(40)H), 4.47–4.50
(m, 1H, C(30)H), 4.98 (dd, J1¼4.4 Hz, J2¼4.2 Hz, 1H,
C(20)H), 6.13 (d, J1¼4.4 Hz, 1H, C(10)H), 8.21 (s, 1H,
C(2)), 8.40 (s, 1H, C(8)); 13C NMR (500 MHz, CDCl3) dC
64.07 (C-50), 70.40 (d, JP,C¼154.3 Hz, PCH2), 80.82 (C-
40), 86.76 (C-20), 88.24 (d, JP,C¼10.8 Hz, C-30), 91.03
(C-10), 121.40 (C-5), 143.37 (C-8), 151.49 (C-6), 155.49
(C-2), 158.26 (C-4); 31P NMR (500 MHz, D2O) dP 14.52;
Exact mass calcd for C11H16N5O7PNa [M+Na]+ 384.0685,
found 384.0685.
1H NMR (500 MHz, CDCl3) dH 1.29–1.32 (m, 12H, CH3),
4.05 (dd, J1¼9.5 Hz, J2¼13.9 Hz, 1H, PCHa), 4.12 (dd,
J1¼8.1 Hz, J2¼13.9 Hz, 1H, PCHb), 4.58 (m, 6H, C(30)H,
C(40)H, C(50)H2, CH(CH3)2), 6.11 (s, 1H, C(20)H), 6.67 (s,
1H, C(10)H), 7.30–7.67 (m, 9H, Ar-H), 8.02–8.06 (m, 6H,
Ar-H), 8.50 (s, 1H, C(8)H), 8.79 (s, 1H, C(2)H), 9.55 (s,
1H, NH); 13C NMR (500 MHz, CDCl3) dC 23.79 (CH3),
64.17 (C-50), 65.05 (d, JP,C¼168 Hz, PCH2), 71.32 (d,
JP,C¼3.9 Hz, CH(CH3)2), 71.36 (d, JP,C¼3.9 Hz,
CH(CH3)2), 71.51 (CH(CH3)2), 80.54 (C-20), 83.42 (C-40),
85.38 (d, JP,C¼9.8 Hz, C-30), 88.00 (C-10), 122.79 (C-5),
128.14, 128.22, 128.52, 129.14, 129.52, 129.71, 132.44,
133.13, 133.55, 133.83 (Ar-C), 141.57 (C-8), 149.62 (C-
6), 151.53 (C-4), 152.70 (C-2), 164.67 (NHC]O), 165.12
(C(20)OC]O), 165.90 (C(50)OC]O); Exact mass calcd
for C38H41N5O10P [M+H]+ 758.2591, found 758.2583.
4.1.28. 1-(Adenin-9-yl)-3-O-(diisopropylphosphono-
methyl)-5-O-methylsulfonyl-a-L-arabinofuranose (33).
To a solution of 32 (128 mg, 0.29 mmol) in 200 mL of pyr-
idine was added dropwise MsCl (0.35 mmol) at 0 ꢀC. The
reaction mixture was warmed to room temperature and
stirred overnight. The reaction mixture was concentrated
and coevaporated with 5 mL toluene two times in vacuo.
The residue was partitioned between H2O (20 mL) and
DCM (3ꢂ50 mL). The combined organic layer was
washed with brine, dried over Na2SO4, and concentrated
in vacuo. The residue was purified by chromatography on
a silica gel column (DCM/MeOH¼15:1) to afford 33
(70 mg, 0.14 mmol) as a colorless amorphous solid in
48% yield.
4.1.26. 1-(Adenin-9-yl)-3-O-(diisopropylphosphono-
methyl)-a-L-arabinofuranose (32). A solution of 31

D. Vin˜a et al. / Tetrahedron 63 (2007) 2634–2646
2645
1H NMR (200 MHz, CDCl3) dH 1.31–1.37 (m, 12H,
C(CH3)2), 3.10 (s, 3H, CH3), 3.91 (dd, J1¼14.4 Hz, J2¼
5.6 Hz, 1H, PCHa), 4.06–4.21 (m, 2H, PCHb, C(40)H),
4.43 (dd, J1¼11.7 Hz, J2¼4.0 Hz, 1H, C(50)Ha), 4.50 (dd,
J1¼11.7 Hz, J2¼2.9 Hz, 1H, C(50)Hb), 4.64–4.83 (m, 3H,
CH(CH3)2, C(30)H), 5.16 (dd, J1¼5.1 Hz, J2¼5.9 Hz, 1H,
C(20)H), 5.91–5.94 (m, 2H, NH2, C(10)H), 7.93 (s, 1H,
C(8)H), 8.27 (s, 1H, C(2)H); 13C NMR (200 MHz, CDCl3)
88.69 (C-10), 120.62 (C-5), 144.21 (C-8), 150.85 (C-6),
155.07 (C-2), 157.86 (C-4); 31P NMR (500 MHz, D2O) dP
13.88; Exact mass calcd for C11H15N5O6P [M+H]+
344.0760, found 344.0753.
4.1.31. 1-(Adenin-9-yl)-3-O-(diphosphorylphosphono-
methyl)-2,5-anhydro-a-L-arabinofuranose triethylam-
monium salt (35). After conversion of the sodium salt of
compound 5 into the triethylammonium form on a Sepha-
dex-DEAE ion exchange column (from 0 to 0.5 M TEAB
in 45 min) the phosphonate nucleoside was lyophilized
overnight. To the dried product (10 mg, 22 mmol), dry DMF
(2 mL) and N,N0-carbonyldiimidazole (36 mg, 220 mmol)
were added under anhydrous conditions. The reaction mix-
ture was stirred under nitrogen for 4 h and followed up by
TLC (i-PrOH/NH4OH concd/H2O¼6:3:1; 0.1 M NH4HCO3/
i-PrOH¼2:8). After quenching the reaction with methanol
(198 mmol, 8 mL) for 30 min to cleave the excess of N,N0-car-
bonyldiimidazole, tris(tetra-n-butylammonium) hydrogen
pyrophosphate (0.4 M solution in dry DMF, 270 mmol,
675 mL) was added. The solution was stirred for 36 h at
room temperature after which it was quenched by evapora-
tion of the solvent under reduced pressure. Analytical ion
exchange chromatography on a mono-Q column (1 M
TEAB from 0 to 50% in 30 min) showed a double peak at
the diphosphorylphosphonate nucleoside position. The dried
reaction mixture was redissolved in 1 M TEAB (1 mL) and
1 N NaOH (44 mmol, 44 mL) was added. The reaction
mixture was stirred for 30 min after the addition of NaOH
and subsequently concentrated to dryness under reduced
pressure. Purification of the diphosphate analog of the phos-
phonate nucleosidewas performedon an ion exchange mono-
Q column (1 M TEAB from 0 to 50% in 30 min). Compound
35 was obtained with a yield of 10% (2 mmol, 1.2 mg).
dC 23.85 (CH3), 23.94 (CH3), 37.72 (SCH3), 66.41 (d, JP,C
¼
167.9 Hz, PCH2), 67.53 (C-50), 71.62 (d, JP,C¼6.1 Hz,
POCH), 72.12 (d, JP,C¼6.1 Hz, POCH), 78.82 (C-40),
79.97 (C-20), 86.51 (d, JP,C¼4.6 Hz, C-30), 89.72 (C-10),
120.25 (C-5), 139.71 (C-8), 149.51 (C-6), 152.91 (C-2),
155.58 (C-4); Exact mass calcd for C18H31N5O9PS
[M+H]+ 524.1580, found 524.1573.
4.1.29. 1-(Adenin-9-yl)-3-O-(diisopropylphosphono-
methyl)-2,5-anhydro-a-L-arabinofuranose (34). Toasolu-
tion of 33 (65 mg, 0.12 mmol) in dried THF (25 mL) was
added sodium hydride (80% dispersion in mineral oil,
5 mg, 0.16 mmol) at ꢁ78 ꢀC. Then the reaction mixture was
slowly warmed to room temperature. The reaction was
quenched with satd NaHCO3 and concentrated. The residue
was partitioned between H2O (5 mL) and EtOAc (50 mL).
The organic layer was washed with water and brine, dried
over Na2SO4, and concentrated invacuo. The residue was pu-
rified by chromatography on a silica gel column (DCM/
MeOH¼15:1) to afford 34 (44 mg, 0.10 mmol, 83%) as
colorless oil.
1H NMR (200 MHz, CDCl3) dH 1.28–1.32 (m, 12H, CH3),
3.61 (dd, 1H, J1¼14.1 Hz, J2¼8.0 Hz, PCHa), 3.65 (dd,
1H, J1¼14.1 Hz, J2¼8.0 Hz, PCHb), 4.02 (d, J¼8.8 Hz,
1H, C(50)Ha), 4.05 (d, J¼8.8 Hz, 1H, C(50)Hb), 4.47 (br s,
1H, C(40)H), 4.65–4.73 (m, 3H, POCH, C(30)H), 4.84 (s,
1H, C(20)H), 6.18 (s, 1H, C(10)H), 8.20 (s, 1H, C(8)), 8.35
(s, 1H, C(2)); 13C NMR (200 MHz, CDCl3) dC 23.88
(CH3), 64.42 (d, JP,C¼167.5 Hz, PCH2), 71.50 (POCH),
72.05 (C-50), 76.22 (C-40), 77.92 (C-20), 81.83 (C-30),
86.67 (C-10), 119.00 (C-5), 140.06 (C-8), 149.51 (C-4),
152.97 (C-2), 155.19 (C-6); Exact mass calcd for
C17H27N5O6PNa [M+Na]+ 450.1518, found 450.1526.
4.2. Incorporation of phosphonate nucleoside 5 by HIV
reverse transcriptase in a DNA duplex
The incorporation of the modified nucleotide by HIV reverse
transcriptase was probed by addition of the building block to
a mixture containing a 32P labeled primer/template complex
(P1T1, Fig. 4) and enzyme. Incorporation was investigated
by taking aliquots at specific time points and analysis of
the samples by PAGE, autoradiography, and phosphor imag-
ing. Reactions were run for 60 min at 37 ꢀC. Aliquots were
taken after 2, 4, 6, 8, 10, 15, 30, and 60 min and quenched by
addition of a double volume of a stopmix (90% v/v form-
amide, 0.05% w/v bromophenol blue, 0.05% w/v xylene
cyanol blue, 50 mM EDTA) and heating of this mixture at
90 ꢀC for 5 min. The samples were analyzed by electropho-
resis on a 16% denaturing polyacrylamide gel with TBE
(89 mM Tris-borate, 2 mM EDTA, pH 8.3) as a buffer. The
amount of the elongated primer was quantified using
Optiquant image analysis software (Packard).
4.1.30. 1-(Adenin-9-yl)-3-O-(phosphonomethyl)-2,5-an-
hydro-a-L-arabinofuranose disodium salt (5). To a solu-
tion of 34 (40 mg, 0.09 mmol) and Et3N (1 mL) in DCM
(25 mL) was added iodotrimethylsilane (0.12 mL,
0.90 mmol) at 0 ꢀC. The reaction mixture was stirred for
2 h. The reaction was quenched with 1.0 M TEAB solution.
The mixture was concentrated and the residue was purified
by column chromatography (CH2Cl2/MeOH¼2:1, 1:1, 1:2)
to give crude 5 as triethylammonium salt. Purification using
Sephadex-DEAE A-25 with gradient TEAB solution from
0.01 to 0.5 M and ion exchanges by the Dowex-Na+ resin
afforded 5 (15 mg, 0.04 mmol) as a colorless solid after
precipitation from diethylether in 44% yield. 1H NMR
(200 MHz, CDCl3) dH 3.42–3.46 (m, 1H, C(50)Ha), 3.56–
3.60 (m, 1H, C(50)Hb), 4.06 (d, J¼8.9 Hz, 1H, PCHa), 4.14
(d, J¼8.9 Hz, 1H, PCHb), 4.56 (s, 1H, C(40)H), 4.83 (s,
1H, C(30)H), 5.05 (s, 1H, C(20)H), 6.05 (s, 1H, C(10)H),
8.12 (s, 1H, C(8)H), 8.36 (s, 1H, C(2)H); 13C NMR
(200 MHz, CDCl3) dC 69.62 (d, JP,C¼153.4 Hz, PCH2),
74.87 (C-50), 78.68 (C-40), 81.10 (C-20), 83.55 (C-30),
The experiments were carried out so that the primer/
template complex concentration was kept constant at
P1
T1
5’ – CAGGAAACAGCTATGAC- 3’
5’ – CCCCTGTCATAGCTGTTTCCTG- 3’
Figure 4. Primer and template sequences used in single nucleotide incorpo-
ration assay for adenine base nucleotides.

2646
D. Vin˜a et al. / Tetrahedron 63 (2007) 2634–2646
250 nM while the diphosphorylphosphonate nucleoside and
the enzyme concentrations were varied.
References and notes
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The authors thank Gilead Sciences for financial support. The
authors thank C. Biernaux for excellent editorial help.
ꢁ
Dr. D.V. is grateful to Xunta de Galicia, Consellerıa de
ꢁ
Innovacion, Industria e Comercio, for a fellowship.
14. Singh, S. K.; Nielsen, P.; Koshkin, A. A.; Wengel, J. Chem.
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